Regulation of vacuolar V-ATPase activity and Dynamic control of vacuolar functions

液泡V-ATP酶活性的调节和液泡功能的动态控制

• 批准号: 10219206
• 负责人: ANRAKU Yasuhiro
• 金额: $ 20.93万
• 依托单位: Teikyo University of Science and Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research on Priority Areas
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-10219206/
• 关键词: Yeast; Vacuole; V-ATPase; VMA3 gene; VIS1 gene; FK506; VMA14遺伝子; VMA15遺伝子

Y. Anraku et al. found a novel ATPase on the vacuolar membrane of Saccharomyces cerevisiae in 1980 It functions as a major proton pump of the vacuole. The enzyme is a supra-macromolecular complex consisting of at least 13 subunits. Its physiological functions are proposed to participate in dynamic controls of vacuolar functions, particularly the regulation of intracellular calcium ion and proton concentrations. In this project, we intended to study a comprehensive view on the regulation of the V-ATPase activity and vacuolar dynamics in cell physiology.The results are : 1) Applying a newly established method of whole vacuolar patch clamp. we determined an ATP-dependent inward current of 100 pA across the vacuolar membrane with a proton/ATP ratio of 3.5. This provides direct evidence that the enzyme acts as a proton pump under physiological conditions 2) The structure and function of novel genes, VMA14 and VMA15 were determined. The products have a regulatory role for the assembly of subunits of the V-ATPase on the vacuolar membrane. 3) A novel gene, VIS1 was isolated and its structure was determined. The product (214 amino acid residues) is a membrane protein enbedded in the vacuole. Its function relates to a regulatory system for Ca^<2+>-sensitive growth with a cross interaction of the UBP signaling network.

Y. Anraku等人于1980年在酿酒酵母的液泡膜上发现了一种新型ATP酶，其功能是液泡的主要质子泵。该酶是由至少13个亚基组成的超大分子复合物。它的生理功能被认为参与液泡功能的动态控制，特别是细胞内钙离子和质子浓度的调节。本课题旨在从细胞生理学的角度对V-ATP酶活性和空泡动力学的调节进行全面的研究，主要结果如下：1）应用新建立的全空泡膜片钳技术。我们测定了质子/ATP比为3.5时，跨液泡膜的ATP依赖性内向电流为100 pA。这为该酶在生理条件下充当质子泵提供了直接证据。2）确定了新基因VMA 14和VMA 15的结构和功能。该产物对液泡膜上V-ATPase亚基的组装具有调节作用。3)分离到一个新基因VIS 1，并对其结构进行了鉴定。该产物（214个氨基酸残基）是包埋在液泡中的膜蛋白。它的功能与钙敏感性生长的调节系统和UBP信号网络的交叉相互作用有关。

项目成果

Yabe, I., Horiuchi, K., Nakahara, K, Hiyama, Y., Yamanaka, Y., 他: "Preparation and Utilization of a Giant Cell Containing a Giant Vacuole"J.Biol.Chem.. 274. 34903-34910 (1999)

Yabe, I.、Horiuchi, K.、Nakahara, K、Hiyama, Y.、Yamanaka, Y.等人：“含有巨液泡的巨细胞的制备和利用”J.Biol.Chem.. 274。 34903- 34910 (1999)

Nakajima-Shimada, J., Sakaguchi, S., Tsuji, F.I., Anraku, Y., Iida, H: "Ca^<2+> signal is generated only once in the mating pheromone response pathway in Saccharomyces cerevisiae"Cell Struct. Funct.. 25. 125-131 (2000)

Nakajima-Shimada，J.，Sakaguchi，S.，Tsuji，F.I.，Anraku，Y.，Iida，H：“Ca ^ 2 >信号在酿酒酵母的交配信息素响应途径中仅产生一次”细胞结构。

Yabe, I.: "Preparation and Utilization of a Giant Cell Containing a Giant Vacuole"J. Biol. Chem.. 274. 34903-34910 (1999)

Yabe，I.：“含有巨液泡的巨细胞的制备和利用”J。

Nakajima-Shimada, J., Sakaguchi, S., Tsuji, F.I., Anraku, Y., Iida, H H: "Ca^<2+> signal is generated only once in the mating pheromone response pathway in Saccharomyces cerevisiae"Cell Struct. Fun. 25. 125-131 (2000)

Nakajima-Shimada，J.，Sakaguchi，S.，Tsuji，F.I.，Anraku，Y.，Iida，H H：“Ca ^ 2 >信号在酿酒酵母的交配信息素响应途径中仅产生一次”细胞结构。

Nakajima-Shimada,J.,Sakaguchi,S.,Tsuji,F.I.,Anraku,Y.,Iida,H: "Ca^<2+> signal is generated only once in the mating pheromone response pathway in Saccharomyces cerevisiae,"Cell Struct.Funct.. 25. 125-131 (2000)

Nakajima-Shimada，J.，Sakaguchi，S.，Tsuji，F.I.，Anraku，Y.，Iida，H：“Ca ^ 2 >信号在酿酒酵母的交配信息素响应途径中仅产生一次，”细胞结构。